Development of Drugs that Modify CNS Innate Immunity for the Treatment of Multiple Sclerosis

项目总结 中枢神经系统(CNS)中常驻细胞(如星形胶质细胞和小胶质细胞)驱动的病理学 在多发性硬化症(MS)中的重要作用，特别是在其最终实现的进展期 大多数多发性硬化症患者。然而，目前还没有治疗星形胶质细胞和小胶质细胞致病的药物。 活动。我们开发了一种体外实验来鉴定调节星形胶质细胞致病活性的化合物。 这些研究确定了化合物A-38，一种产生促红细胞生成素的肝细胞的抑制物 肿瘤受体B3(EphB3)作为星形胶质细胞致病活性的抑制因子，因此，作为一种 治疗进展性多发性硬化(PMS)的先导治疗化合物。此外，我们的数据表明，小胶质细胞驱动的 星形胶质细胞中的EphB3信号促进中枢神经系统炎症和神经变性。肾上腺素受体是 已知参与中枢神经系统发育，然而我们的发现定义了EphB3在中枢神经系统中的新角色 并将其确定为治疗干预的潜在靶点。我们假设EphB3 抑制激酶将为经前综合症提供一种有效的治疗方法，并可能 神经系统疾病。因此，我们建议开发新的A-38类似物来治疗经前综合症。我们的具体目标是： 特异性目标1：EPHB3激酶抑制剂的优化和体外评价。我们会 设计、合成和评估多达150个A-38类似物的性能。这些研究的目标是 优化A-38新类似物的效力和激酶选择性，同时保持良好的类药物特性 以及口服分娩后的脑部暴露。在该项目的R61阶段，我们将集成迭代 构效关系(SAR)研究和基于结构的药物设计优化A-38效力和 激酶选择性，同时保持良好的类药物特性。我们将首先评估针对EphB3的A-38类似物 在EphB2、EphB4和EphA4激酶检测中的活性和选择性。前景看好的化合物将发展到 表型分析用于识别那些调节星形胶质细胞致病活动的分子。化合物 然后将在类药物性质分析中表征兴趣，以选择用于PO PK研究的领先化合物。 在这个项目的R61阶段结束时，我们希望确定一个主要的EphB3抑制剂和1-2个后备 适用于进入R33阶段的体内疗效测试。 特定目的2：先导化合物(S)在经前综合征临床前模型中的评价。在.期间 该项目的R33阶段，我们将评估NOD EAE中的主要EphB3抑制剂和1-2个后备 铜酮诱导的小鼠临床前模型，概括了经前综合征的几个方面。我们将评估 这些化合物对疾病发展、中枢神经系统炎症、轴突丢失和脱髓鞘的影响，如 通过组织病理学、流式细胞术和基因表达分析确定。 综上所述，该项目将确定一种新的EphB3抑制剂，具有适合开发的类药物特性 作为治疗经前综合症和其他神经疾病的候选药物，获得美国国立卫生研究院和/或行业的支持。

PROJECT SUMMARY Pathology driven by resident cells in the central nervous system (CNS) such as astrocytes and microglia plays an important role in multiple sclerosis (MS), particularly for its progressive stage which is eventually achieved by most MS patients. However, no drugs are available for the modulation astrocyte and microglia pathogenic activities. We developed an in vitro assay to identify compounds that modulate astrocyte pathogenic activities. These studies identified compound A-38, an inhibitor of the Erythropoietin-producing hepatocellular carcinoma receptor B3 (EphB3), as a suppressor of astrocyte pathogenic activities and, consequently, as a lead therapeutic compound for progressive MS (PMS). Moreover, our data suggest that microglia-driven EphB3 signaling in astrocytes promotes CNS inflammation and neurodegeneration. Ephrin receptors are known to participate in CNS development, however our findings define a novel role for EphB3 in CNS inflammation and identify it as potential target for therapeutic intervention. We hypothesize that EphB3 kinase inhibition will provide an efficacious therapeutic approach for PMS, and potentially other neurologic diseases. Thus, we propose to develop new A-38 analogs to treat PMS. Our Specific Aims are: SPECIFIC AIM 1: OPTIMIZATION AND IN VITRO EVALUATION OF EPHB3 KINASE INHIBITORS. We will design, synthesize and evaluate the properties of up to 150 analogs of A-38. The goal of these studies is to optimize the potency and kinase selectivity of new analogs of A-38, while maintaining good drug-like properties and brain exposure following oral delivery. During the R61 phase of this project, we will integrate iterative structure-activity relationship (SAR) studies and structure based drug design to optimize A-38 potency and kinase selectivity, while maintaining good drug-like properties. We will first evaluate A-38 analogs for EphB3 activity and for selectivity in EphB2 EphB4, and EphA4 kinase assays. Promising compounds will advance to phenotypic assays to identify those molecules that modulate astrocyte pathogenic activities. Compounds of interest will then be characterized in drug-like property assays to select leading compounds for PO PK studies. At the end of the R61 phase of this project we expect to identify a lead EphB3 inhibitor and 1-2 back-ups suitable for advancement into in vivo efficacy testing in the R33 phase. SPECIFIC AIM 2: EVALUATION OF LEAD COMPOUND(S) IN PMS PRE-CLINICAL MODELS. During the R33 phase of this project, we will evaluate the lead EphB3 inhibitor and 1-2 back-ups in the NOD EAE and cuprizone-induced murine pre-clinical models which recapitulate several aspects of PMS. We will evaluate the effects of the compounds on disease development, CNS inflammation, axonal loss and demyelination, as determined by histopathology, flow cytometry and gene expression analyses. In summary, this project will identify a novel EphB3 inhibitor with drug-like properties suitable for development as a candidate therapeutic for PMS and other neurologic diseases with NIH and/or industry support.